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Marc Fortin,1 Del R Dorscheid,2 Moishe Liberman,3 Simon Martel,1 Tawimas Shaipanich2 1Department of Pulmonary Medicine and Thoracic Surgery, University of Laval Institut Universitaire de Cardiologie et de Pneumologie de Québec (IUCPQ), Quebec City, Quebec, Canada; 2Department of Medicine, St. Paul’s Hospital, University of British Columbia, Vancouver, British Columbia, Canada; 3Division of Thoracic Surgery, Centre hospitalier de l’Université de Montréal (CHUM), Montreal, Quebec, CanadaCorrespondence: Marc Fortin, Department of Pulmonary Medicine and Thoracic Surgery, Quebec Heart and Lung Institute, Laval University, 2725 Chemin Sainte-Foy, Quebec City, QC, G1V 4G5, Canada, Tel +1 418-656-8711, Email marc.fortin@criucpq.ulaval.caPurpose: Chronic bronchitis (CB), a chronic obstructive pulmonary disease (COPD) phenotype defined by persistent mucus hypersecretion and cough, is associated with poor quality of life, exacerbations, and lung function impairment. Bronchial Rheoplasty (BR) delivers non-thermal pulsed electric fields to airway epithelium and submucosa. Preliminary studies demonstrated reduced airway goblet cell hyperplasia and symptom improvement in response to BR. This study aimed to further assess the safety and clinical feasibility of BR in the setting of CB.Patients and Methods: This 3-center, single-arm study evaluated the safety and feasibility of BR in Canadian patients. The major inclusion criteria were the sum of CAT first 2 questions (cough and mucus) ≥ 7 out of 10 and FEV1 ≥ 30% predicted. Right-sided airways were treated first; left, 1 month later. Serious adverse events (SAEs) were tabulated through 12 months. Outcomes were evaluated using the SGRQ and CAT.Results: Ten patients with CB were enrolled and followed for 12 months. The BR procedure was successful in all patients (mean age 69 ± 5.8 years, post-BD FEV1 77.1 ± 28.3, SGRQ 56.2 ± 8.8, CAT 25.4 ± 4.7). Only one SAE, a COPD exacerbation 13 days following the BR procedure, was considered device related. No additional SAEs occurred through 12 months, and 90% of the patients were CAT responders (≥ 2-point improvement) at 3 and 6 months. Similar results were observed in SGRQ.Conclusion: BR was safe and well-tolerated. Meaningful symptom improvement was observed through 12 months, suggesting BR may be a viable treatment option for patients with CB.Keywords: chronic bronchitis, COPD, bronchoscopy, pulsed electric fields

A simple method for rapid and large scale (over 109 cells) separation of peritoneal macrophages by incubation in a glass-bead column and elution with buffered ethylenediamine tetraacetic acid (EDTA) solution is described. Macrophages obtained were about 90 per cent pure with the recovery rates of 38-53 per cent depending upon the sample size, and 83 per cent viability. Lymphocyte contamination was reduced to 5 per cent or less. [ABSTRACT FROM AUTHOR]

To elucidate the important cellular and molecular drivers of pulmonary long COVID, we generated a single-cell transcriptomic map of the airway mucosa using bronchial brushings from patients with long COVID who reported persistent pulmonary symptoms.Adults with and without long COVID were recruited from the general community in Greater Vancouver, Canada. The cohort was divided into those with pulmonary long COVID (PLC), which was defined as persons with new or worsening respiratory symptoms following at least one year from their initial acute SARS-CoV-2 infection (N=9); and control subjects defined as SARS-CoV-2 infected persons whose acute respiratory symptoms had fully resolved or individuals who had no history of acute COVID-19 (N=9). These participants underwent bronchoscopy from which a single cell suspension was created from bronchial brush samples and then sequenced.A total of 56 906 cells were recovered for the downstream analysis, with 34 840 cells belonging to the PLC group, which strikingly showed a unique cluster of neutrophils in the PLC group (p<0.05). Ingenuity Pathway Analysis revealed that the neutrophil degranulation pathway was enriched across epithelial cell clusters. Differential gene expression analysis between the PLC and control groups demonstrated upregulation of inflammatory chemokines and epithelial barrier dysfunction across epithelial cell clusters, as well as over-expression of mucin genes across secretory cell clusters.In conclusion, a single-cell transcriptomic landscape of the small airways suggest that neutrophils may play a significant role in mediating the chronic small airway inflammation driving pulmonary symptoms of long COVID., (Copyright ©The authors 2024.)

Background: DNA methylation may be a link between HIV, aging, and the increased risk of lung comorbidities. We investigated whether bronchoalveolar lavage (BAL) cells of people living with HIV (PLWH) demonstrate epigenetic disruptions and advanced epigenetic aging., Methods: BAL cell DNA methylation from 25 PLWH and 16 HIV-uninfected individuals were tested for differential methylation of Alu and LINE-1 sites, markers of aging. We used a weighted gene correlation network analysis to identify HIV- and age-associated co-methylation networks. We tested the effect of HIV on DNA methylation using a robust linear model (false discovery rate < 0.10)., Results: The BAL cells of PLWH were marked by global hypomethylation in both Alu and LINE-1 elements. Six co-methylated CpG networks were identified that were significantly associated with age; of these, the red module was significantly differentially methylated in PLWH and enriched pathways (e.g., Ras signaling and T-cell receptors). We identified 6428 CpG sites associated with HIV., Conclusions: We have shown here for the first time that alterations in the DNA methylation of BAL cells in the lung with HIV show a pattern of advanced aging. This study strongly supports that HIV may contribute to an increased the risk of lung comorbidities through the epigenetics of aging.

Single-cell RNA sequencing (scRNA-seq) is an important tool for understanding disease pathophysiology, including airway diseases. Currently, the majority of scRNA-seq studies in airway diseases have used invasive methods (airway biopsy, surgical resection), which carry inherent risks and thus present a major limitation to scRNA-seq investigation of airway pathobiology. Bronchial brushing, where the airway mucosa is sampled using a cytological brush, is a viable, less invasive method of obtaining airway cells for scRNA-seq. Here we describe the development of a rapid and minimal handling protocol for preparing single-cell suspensions from bronchial brush specimens for scRNA-seq. Our optimized protocol maximizes cell recovery and cell quality and facilitates large-scale profiling of the airway transcriptome at single-cell resolution.

Background: People living with HIV (PLWH) are at increased risk of developing Chronic Obstructive Pulmonary Disease (COPD) independent of cigarette smoking. We hypothesized that dysbiosis in PLWH is associated with epigenetic and transcriptomic disruptions in the airway epithelium., Methods: Airway epithelial brushings were collected from 18 COPD + HIV + , 16 COPD - HIV + , 22 COPD + HIV - and 20 COPD - HIV - subjects. The microbiome, methylome, and transcriptome were profiled using 16S sequencing, Illumina Infinium Methylation EPIC chip, and RNA sequencing, respectively. Multi 'omic integration was performed using Data Integration Analysis for Biomarker discovery using Latent cOmponents. A correlation > 0.7 was used to identify key interactions between the 'omes., Results: The COPD + HIV -, COPD -HIV + , and COPD + HIV + groups had reduced Shannon Diversity (p = 0.004, p = 0.023, and p = 5.5e-06, respectively) compared to individuals with neither COPD nor HIV, with the COPD + HIV + group demonstrating the most reduced diversity. Microbial communities were significantly different between the four groups (p = 0.001). Multi 'omic integration identified correlations between Bacteroidetes Prevotella, genes FUZ, FASTKD3, and ACVR1B, and epigenetic features CpG-FUZ and CpG-PHLDB3., Conclusion: PLWH with COPD manifest decreased diversity and altered microbial communities in their airway epithelial microbiome. The reduction in Prevotella in this group was linked with epigenetic and transcriptomic disruptions in host genes including FUZ, FASTKD3, and ACVR1B., (© 2023. The Author(s).)

Epigenetic modifications are common in chronic obstructive pulmonary disease (COPD); however, their clinical relevance is largely unknown. We hypothesized that epigenetic disruptions are associated with symptoms and health status in COPD. We profiled the blood ( n = 57) and airways ( n = 62) of COPD patients for DNA methylation ( n = 55 paired). The patients' health status was assessed using the St. George's Respiratory Questionnaire (SGRQ). We conducted differential methylation analyses and identified pathways characterized by epigenetic disruptions associated with SGRQ scores and its individual domains. 29,211 and 5044 differentially methylated positions (DMPs) were associated with total SGRQ scores in blood and airway samples, respectively. The activity, impact, and symptom domains were associated with 9161, 25,689 and 17,293 DMPs in blood, respectively; and 4674, 3730 and 5063 DMPs in airways, respectively. There was a substantial overlap of DMPs between airway and blood. DMPs were enriched for pathways related to common co-morbidities of COPD (e.g., ageing, cancer and neurological) in both tissues. Health status in COPD is associated with airway and systemic epigenetic changes especially in pathways related to co-morbidities of COPD. There are more blood DMPs than in the airways suggesting that blood epigenome is a promising source to discover biomarkers for clinical outcomes in COPD.

One key feature of Chronic Obstructive Pulmonary Disease (COPD) is that its prevalence increases exponentially with age. DNA methylation clocks have become powerful biomarkers to detect accelerated aging in a variety of diseases and can help prognose outcomes in severe COPD. This study investigated which DNA methylation clock could best reflect airway epigenetic age when used in more accessible blood samples. Our analyses showed that out of six DNA methylation clocks investigated, DNAmGrimAge demonstrated the strongest correlation and the smallest difference between the airway epithelium and blood. Our findings suggests that blood DNAmGrimAge accurately reflects airway epigenetic age of individuals and that its elevation is highly associated with COPD.

The lung is an understudied site of HIV persistence. We isolated 898 subgenomic proviral sequences (nef) by single-genome approaches from blood and lung from nine individuals on long-term suppressive antiretroviral therapy (ART), and characterized genetic diversity and compartmentalization using formal tests. Consistent with clonal expansion as a driver of HIV persistence, identical sequences comprised between 8% to 86% of within-host datasets, though their location (blood vs. lung) followed no consistent pattern. The majority (77%) of participants harboured at least one sequence shared across blood and lung, supporting the migration of clonally-expanded cells between sites. The extent of blood proviral diversity on ART was also a strong indicator of diversity in lung (Spearman's ρ = 0.98, p<0.0001). For three participants, insufficient lung sequences were recovered to reliably investigate genetic compartmentalization. Of the remainder, only two participants showed statistically significant support for compartmentalization when analysis was restricted to distinct proviruses per site, and the extent of compartmentalization was modest in both cases. When all within-host sequences (including duplicates) were considered, the number of compartmentalized datasets increased to four. Thus, while a subset of individuals harbour somewhat distinctive proviral populations in blood and lung, this can simply be due to unequal distributions of clonally-expanded sequences. For two participants, on-ART proviruses were also phylogenetically analyzed in context of plasma HIV RNA populations sampled up to 18 years prior, including pre-ART and during previous treatment interruptions. In both participants, on-ART proviruses represented the most ancestral sequences sampled within-host, confirming that HIV sequences can persist in the body for decades. This analysis also revealed evidence of re-seeding of the reservoir during treatment interruptions. Results highlight the genetic complexity of proviruses persisting in lung and blood during ART, and the uniqueness of each individual's proviral composition. Personalized HIV remission and cure strategies may be needed to overcome these challenges., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2022 Shahid et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Age-related comorbidities such as chronic obstructive pulmonary disease (COPD) are common in people living with human immunodeficiency virus (PLWH). We investigated the relationship between COPD and the epigenetic age of the airway epithelium and peripheral blood of PLWH., Methods: Airway epithelial brushings from 34 PLWH enrolled in the St. Paul's Hospital HIV Bronchoscopy cohort and peripheral blood from 378 PLWH enrolled in The Strategic Timing of Antiretroviral Treatment (START) study were profiled for DNA methylation. The DNA methylation biomarker of age and healthspan, GrimAge, was calculated in both tissue compartments. We tested the association of GrimAge with COPD in the airway epithelium and airflow obstruction as defined by an FEV 1 /FVC<0.70, and FEV 1 decline over 6 years in blood., Findings: The airway epithelium of PLWH with COPD was associated with greater GrimAge residuals compared to PLWH without COPD (Beta=3.18, 95%CI=1.06-5.31, P=0.005). In blood, FEV 1 /FVC1 decline was inversely correlated with GrimAge residuals in blood (r=-0.13, P=0.012). PLWH who had normal lung function but who subsequently developed an FEV 1 /FVC<0.70 over the course of 6 years had higher GrimAge residuals at baseline (Beta=2.33, 95%CI=0.23-4.44, P=0.031)., Interpretation: GrimAge may reflect lung and systemic epigenetic changes that occur with advanced airflow obstruction and may help to identify PLWH with a higher risk of developing COPD., Funding: Canadian Institutes of Health Research and the British Columbia Lung Association. The START substudy was funded by NIH grants: UM1-AI068641, UM1-AI120197, and RO1HL096453., Competing Interests: Declaration of interests SH reports consulting fees from the non-profit organization, Epigenetic Clock Development Foundation, and royalty income from patents surrounding epigenetic clocks. HK reports consulting fees and payments from AbbVie, Janssen, MSD, ViiV, Gilead, and Intercept, outside of the submitted work. KK reports consulting fees and payments from Allergan and Nuvaira, outside of the submitted work. DS reports payments from AstraZeneca, GSK, and Boehringer Ingelheim, outside of the submitted work. MH reports consulting fees and payments from Gilead, Merck, and ViiV, outside of the submitted work. RMN reports payments from ViiV, Gilead, and Theratechnologies, outside of the submitted work. J Montaner reports grants from Gilead, Merck, ViiV, the Public Health Agency of Canada, and the BC Ministry of Health, outside of the submitted work. JML reports a grant from AstraZeneca, outside of the submitted work. The remaining authors report no competing interests., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Rationale: Age-related diseases like chronic obstructive pulmonary disease (COPD) occur at higher rates in people living with human immunodeficiency virus (PLWH) than in uninfected populations. Objectives: To identify whether accelerated aging can be observed in the airways of PLWH with COPD, manifest by a unique DNA methylation signature. Methods: Bronchial epithelial brushings from PLWH with and without COPD and HIV-uninfected adults with and without COPD ( N  = 76) were profiled for DNA methylation and gene expression. We evaluated global Alu and LINE-1 methylation and calculated the epigenetic age using the Horvath clock and the methylation telomere length estimator. To identify genome-wide differential DNA methylation and gene expression associated with HIV and COPD, robust linear models were used followed by an expression quantitative trait methylation (eQTM) analysis. Measurements and Main Results: Epigenetic age acceleration and shorter methylation estimates of telomere length were found in PLWH with COPD compared with PLWH without COPD and uninfected patients with and without COPD. Global hypomethylation was identified in PLWH. We identified 7,970 cytosine bases located next to a guanine base (CpG sites), 293 genes, and 9 expression quantitative trait methylation-gene pairs associated with the interaction between HIV and COPD. Actin binding LIM protein family member 3 ( ABLIM3 ) was one of the novel candidate genes for HIV-associated COPD highlighted by our analysis. Conclusions: Methylation age acceleration is observed in the airway epithelium of PLWH with COPD, a process that may be responsible for the heightened risk of COPD in this population. Their distinct methylation profile, differing from that observed in patients with COPD alone, suggests a unique pathogenesis to HIV-associated COPD. The associations warrant further investigation to establish causality.

The associations between airway eosinophilia, measured in sputum or peripheral blood, and acute exacerbations of chronic obstructive pulmonary disease (AECOPD) are inconsistent. We therefore aimed to determine the association between eosinophilia in bronchoalveolar lavage (BAL) fluid and AECOPD in a clinical cohort. We analyzed differential cell counts from baseline BAL fluid in participants in the DISARM clinical trial (Clinicaltrials.gov #NCT02833480) and classified participants by the presence or absence of BAL eosinophilia (>1% of total leukocytes). We determined the association between BAL eosinophilia and AECOPD over 1 year of follow-up using negative binomial regression and Cox proportional hazards test. N = 63 participants were randomized, and N = 57 had BAL differential cell counts available. Participants with BAL eosinophilia (N = 21) had a significantly increased rate of acute exacerbations (unadjusted incidence rate ratio (IRR) 2.0, p = 0.048; adjusted IRR 2.24, p = 0.04) and a trend toward greater probability of acute exacerbation (unadjusted hazard ratio (HR) 1.74, p = 0.13; adjusted HR 2.3, p = 0.1) in the year of follow-up compared to participants without BAL eosinophilia (N = 36). These associations were not observed for BAL neutrophilia (N = 41 participants), BAL lymphocytosis (N = 27 participants) or peripheral blood eosinophilia at various threshold definitions (2%, N = 37; 3%, N = 27; 4%, N = 16). BAL may therefore be a sensitive marker of eosinophilic inflammation in the distal lung and may be of benefit for risk stratification or biomarker-guided therapy in COPD.

Background: Patients with chronic obstructive pulmonary disease (COPD) are commonly treated with inhaled corticosteroid/long-acting ß2-agonist combination therapy. While previous studies have investigated the host-microbiome interactions in COPD, the effects of specific steroid formulations on this complex cross-talk remain obscure., Methods: We collected and evaluated data from the Study to Investigate the Differential Effects of Inhaled Symbicort and Advair on Lung Microbiota (DISARM), a randomized controlled trial. Bronchoscopy was performed on COPD patients before and after treatment with salmeterol/fluticasone, formoterol/budesonide or formoterol-only. Bronchial brush samples were processed for microbial 16S rRNA gene sequencing and host mRNA sequencing. Longitudinal changes in the microbiome at a community, phylum and genus level were correlated with changes in host gene expression using a Spearman's rank correlation test., Findings: In COPD patients treated with salmeterol/fluticasone, the expression levels of 676 host genes were significantly correlated to changes in the alpha diversity of the small airways. At a genus level, the expression levels of 122 host genes were significantly related to changes in the relative abundance of Haemophilus . Gene enrichment analyses revealed the enrichment of pathways and biological processes related to innate and adaptive immunity and inflammation. None of these changes were evident in patients treated with formoterol/budesonide or formoterol alone., Interpretation: Changes in the microbiome following salmeterol/fluticasone treatment are related to alterations in the host transcriptome in the small airways of patients with COPD. These data may provide insights into why some COPD patients treated with inhaled corticosteroids may be at an increased risk for airway infection, including pneumonia., Funding: The Canadian Institute of Health Research, the British Columbia Lung Association, and an investigator-initiated grant from AstraZeneca.

Background: People with human immunodeficiency virus (PWH) have an increased risk of developing chronic obstructive pulmonary disease (COPD)., Methods: We phenotyped lung macrophages in 4 subgroups-M1 (CD40+CD163-), M2 (CD40-CD163+), double positives (CD40+CD163+), and double negatives and (CD40-CD163-)-and we determined their phagocytic capacity in PWH with and without COPD., Results: People with human immunodeficiency virus with COPD have more double-negative macrophages (84.1%) versus PWH without (54.3%) versus controls (23.9%) (P=.004) and reduced phagocytosis (P=.012). Double-negative macrophages had the worst phagocytic capacity (P<.001)., Conclusions: People with human immunodeficiency virus with COPD have an abundance of nonpolarized macrophages, which have poor phagocytic capacity and therefore predispose PWH to increased risk of disease progression., (© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Rationale: Inhaled corticosteroids (ICS) are commonly prescribed with long-acting β 2 -agonists (LABA) in chronic obstructive pulmonary disease (COPD). To date, the effects of ICS therapy on the airway microbiome in COPD are unknown. Objectives: To determine the effects of ICS/LABA on the airway microbiome of patients with COPD. Methods: Clinically stable patients with COPD were enrolled into a 4-week run-in period during which ICS was discontinued and all participants were placed on formoterol (Form) 12 μg twice daily (BID). The participants were then randomized to budesonide/formoterol (Bud + Form; 400/12 μg BID), fluticasone/salmeterol (Flu + Salm; 250/50 μg BID), or formoterol only (12 μg BID) for 12 weeks. Participants underwent bronchoscopy before and after the 12-week treatment period. The primary endpoint was the comparison of changes in the airway microbiome over the trial period between the ICS/LABA and LABA-only groups. Measurements and Main Results: Sixty-three participants underwent randomization: Bud + Form ( n  = 20), Flu + Salm ( n  = 22), and Form ( n  = 21) groups; 56 subjects completed all visits. After the treatment period, changes in α-diversity were significantly different across groups, especially between Flu + Salm and Form groups (Δrichness: P  = 0.02; ΔShannon index: P  = 0.03). Longitudinal differential abundance analyses revealed more pronounced microbial shifts from baseline in the fluticasone (vs. budesonide or formoterol only) group. Conclusions: Fluticasone-based ICS/LABA therapy modifies the airway microbiome in COPD, leading to a relative reduction in α-diversity and a greater number of bacterial taxa changes. These data may have implications in patients who develop pneumonia on ICS. Clinical trial registered with www.clinicaltrials.gov (NCT02833480).

The classical M1/M2 polarity of macrophages may not be applicable to inflammatory lung diseases including chronic obstructive pulmonary disease (COPD) due to the complex microenvironment in lungs and the plasticity of macrophages. We examined macrophage sub-phenotypes in bronchoalveolar lavage (BAL) fluid in 25 participants with CD40 (a M1 marker) and CD163 (a M2 marker). Of these, we performed RNA-sequencing on each subtype in 10 patients using the Illumina NextSeq 500. Approximately 25% of the macrophages did not harbor classical M1 or M2 surface markers (double negative, DN), and these cells were significantly enriched in COPD patients compared with non-COPD patients (46.7% vs. 14.5%, p < 0.001). 1886 genes were differentially expressed in the DN subtype compared with  all other subtypes at a 10% false discovery rate. The 602 up-regulated genes included 15 mitochondrial genes and were enriched in 86 gene ontology (GO) biological processes including inflammatory responses. Modules associated with cellular functions including oxidative phosphorylation were significantly down-regulated in the DN subtype. Macrophages in the human BAL fluid, which were negative for both M1/M2 surface markers, harbored a gene signature that was pro-inflammatory and suggested dysfunction in cellular homeostasis. These macrophages may contribute to the pathogenesis and manifestations of inflammatory lung diseases such as COPD.

Background: No currently approved intervention counteracts airway metaplasia and mucus hypersecretion of chronic bronchitis in COPD. However, metered cryospray (MCS) delivering liquid nitrogen to the tracheobronchial airways ablates abnormal epithelium and facilitates healthy mucosal regeneration. The objective of this study was to evaluate the feasibility, efficacy and safety of MCS in chronic bronchitis., Methods: Patients with a forced expiratory volume in 1 s of 30-80% predicted who were taking optimal medication were recruited. Primary outcomes were feasibility (completion of treatments), efficacy (3-month change in St George's Respiratory Questionnaire (SGRQ)) and safety (incidence of adverse events). Secondary outcomes were lung function, exercise capacity and additional patient-reported outcomes., Results: 35 patients, 19 male/16 female, aged 47-76 years, Global Initiative for Chronic Obstructive Lung Disease grade I (n=3), II (n=10) and III (n=22), underwent staggered liquid nitrogen treatments to the tracheobronchial tree. 34 patients completed three treatments, each lasting 34.3±12.1 min, separated by 4-6 weeks; one withdrew after the first treatment. ∼1800 doses of MCS were delivered. Clinically meaningful improvements in patient-reported outcomes were observed at 3 months: change in SGRQ -6.4 (95% CI -11.4 to -1.3; p=0.01), COPD Assessment Test (CAT) -3.8 (95% CI -6.4 to -1.3; p<0.01) and Leicester Cough Questionnaire (LCQ) 21.6 (95% CI 7.3 to 35.9; p<0.01). Changes in CAT were durable to 6 months (-3.4, 95% CI -5.9 to -0.9; p=0.01); changes in SGRQ and LCQ were durable to 9 months (-6.9, 95% CI -13.0 to -0.9; p=0.03 and 13.4, 95% CI 2.1 to 24.6; p=0.02, respectively. At 12 months, 14 serious adverse events were recorded in 11 (31.4%) subjects; six (43%) moderate and eight (57%) severe. Nine were respiratory-related: six exacerbations of COPD, two pneumonias and one case of increased coughing; all recovered without sequelae. None were serious device- or procedure-related adverse events., Conclusion: MCS is safe, feasible and associated with clinically meaningful improvements in multidimensional patient-reported outcomes., Competing Interests: Conflict of interest: J.L. Garner has nothing to disclose. Conflict of interest: T. Shaipanich has nothing to disclose. Conflict of interest: J.E. Hartman has nothing to disclose. Conflict of interest: C.M. Orton has nothing to disclose. Conflict of interest: C. Caneja has nothing to disclose. Conflict of interest: K. Klooster has nothing to disclose. Conflict of interest: J. Thornton has nothing to disclose. Conflict of interest: D.D. Sin has received honoraria for speaking engagements from AstraZeneca and Boehringer Ingelheim, and funding for research projects from AstraZeneca, Boehringer Ingelheim and Merck. Conflict of interest: D.J. Slebos reports grants and non-financial support from, and was PI for and advisor to CSA Medical, USA, during the conduct of the study; grants and non-financial support from, and was PI for and advisor to PulmonX, USA, Nuvaira, USA and PneumRx/BTG, USA, and was PI for and advisor to FreeFlowMedical, USA, outside the submitted work. Conflict of interest: P.L Shah was reimbursed by CSA Medical for travel expenses incurred during the training with the medical device and trial protocol development; the hospital was reimbursed for all clinical trial related costs, and reports personal fees from CSA Medical, Boston Scientific, Broncus, Creo Medical, Nuvaira, Olympus, Medtronic and PneumRX/BTG as consultant on scientific advisory board; and sponsorship to Imperial College for a bronchoscopy course from ERBE, Cook medical, Medtronic, Boston Scientific, Broncus, Pulmonx, Olympus and PneumRX/BTG, outside the submitted work., (Copyright ©ERS 2020.)

Background: Transbronchial needle aspiration (TBNA) of peripheral lung nodules can be difficult with conventional devices due to their limited flexibility. A promising new technology for accessing these lesions is the PeriView FLEX TBNA device, which has a flexible spiral-grooved needle. The present study reports the unique cytologic features, diagnostic value, and potential pitfalls of PeriView FLEX TBNA specimens., Methods: This study retrospectively evaluates 113 consecutive cases of lung nodules sampled using the PeriView FLEX device with radial endobronchial ultrasound guidance., Results: PeriView FLEX specimens were satisfactory for evaluation in 111 of 113 cases (98%). A diagnosis of malignancy was made on 64 specimens (57%), with 100% specificity and 70% sensitivity for malignancy. In 4 cases, the PeriView FLEX sample was the only specimen from bronchoscopy that was diagnostic of malignancy. Of the 64 PeriView FLEX specimens with malignant cells, 58 (91%) were adequate for immunohistochemistry and 44 (69%) were adequate for molecular genetic testing. Potential pitfalls were largely ameliorated through education regarding the unique features of PeriView FLEX samples, such as the expected abundance of anthracotic pigment and the paucity of lymphocytes., Conclusions: TBNA using the PeriView FLEX device to sample pulmonary nodules contributed to the diagnostic value of bronchoscopy and tended to provide sufficient tissue for ancillary studies. Many of the possible pitfalls may be avoided through consideration of the unique cytologic features associated with this novel sampling method., (© 2020 American Cancer Society.)

Objectives: The FDA approved PD-L1 tests for anti-PD-L1 immunotherapy are for surgical or histology specimens. It is not clear if cytology specimens could be used for PD-L1 testing to guide immunotherapy. In this study, we assess the suitability of EBUS-FNA cytology specimens for the testing of PD-L1., Materials and Methods: Consecutive patients with Non-small cell lung cancer (NSCLC) underwent EBUS procedure between January 1, 2017 and March 31, 2018 for PD-L1 testing were included. The cell blocks of EBUS-FNA cytology specimens were used for PD-L1 testing using Dako 22C3 phamDx antibody according to the Dako protocol. PD-L1 protein expression in tumor cells is determined by using Tumor Proportion Score (TPS)., Results and Conclusion: Of the 265 EBUS-FNA specimens from 262 patients sent for testing, 230 (86.8%) were adequate for PD-L1 testing. Of the 34 NSCLC patients with both histology and EBUS-FNA cytology specimens tested for PD-L1, the results from different specimen types had a concordance of 91.3%. The PD-L1 results from 16 paired specimens from the same anatomic site had 100% agreement. The rates of PD-L1 TPS ≥ 50% were significantly higher in the metastatic tumors in the lymph nodes than in the lung primary lesions. Therefore, EBUS-FNA cytology specimen is suitable for PD-L1 testing in patients with advanced NSCLC. The metastatic tumors in mediastinal lymph nodes appear to have higher PD-L1 expression than primary lesions., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Lung cancer is associated with high mortality. It can present as one or more pulmonary nodules identified on computed tomography (CT) chest scans. The National Lung Screening Trial has shown that the use of low-dose CT chest screening can reduce deaths due to lung cancer. High adherence to appropriate follow-up of positive results, including imaging or interventional approaches, is an important aspect of pulmonary nodule management. Our study is one of the first to evaluate the current practice in managing pulmonary nodules and to explore potential causes for nonadherence to follow-up. This is a retrospective analysis at St. Paul's Hospital, a tertiary healthcare center in Vancouver, British Columbia, Canada. We first identified CT chest scans between January 1 to June 30, 2014, that demonstrated one or more pulmonary nodules equal to or greater than 6 mm in diameter. We then looked for evidence of interventional (surgical resection or biopsy, or bronchoscopy for transbronchial biopsy and cytology) and radiological follow-up of the pulmonary nodule by searching on the province-wide CareConnect eHealth Viewer patient database. A total of 1614 CT reports were analyzed and 139 (8.6%) had a positive finding. Out of the 97 patients who received follow-up, 54.6% ( N  = 53) was referred for a repeat CT chest scan and 36.1% ( N  = 35) and 9.3% ( N  = 9) were referred for interventional biopsy and surgical resection, respectively. In our study, 30.2% ( N  = 42) of the patients with pulmonary nodules were nonadherent to follow-up. Despite the radiologist's recommendation for follow-up within a certain time interval, only 36% had repeat imaging in a timely manner. Our findings reflect the current practice in the management of pulmonary nodules and suggest that there is a need for improvement at our academic center. Adherence to follow-up is important for the potentially near-future implementation of lung cancer screening.

The management of cancer in the periphery lung is in critical need of new strategies. Here, the development and test of a novel miniature Raman probe capable of navigating the peripheral lung architecture is reported. The probe was 1.35 mm in diameter, with a minimum bend radius of 13 mm and had a large light collection area for its size. Peripheral lung Raman spectra were successfully obtained from normal tissue and cancerous nodule using the probe coupled to a home-made rapid Raman spectroscopy system with a fast integration time of 1 second and a low excitation power of 15 mW. This is the first time in vivo Raman spectra from the periphery lung being reported. The collected spectra showed lipid, protein and deoxyhemoglobin signatures that might be useful for classifying pathology. Large scale clinical study is planned to confirm the utility of this new technology for improving periphery lung cancer detection. Left: Radial ultrasound image of a peripheral lung nodule: size given by crosshairs D1 and D2. Right: Truncated Raman spectra of a cancerous nodule, whole blood, and normal peripheral airway tissue. Spectra were shifted on intensity scale for clarity., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)